Adventitious regeneration in styrian oil pumpkin in relation to the endoreduplication pattern and induced tetraploidy on fusaric acid-supplemented media

Abstract

We attempted to improve the adventitious regeneration protocol in styrian oil pumpkin for subsequent use in somaclonal variation studies. The endoreduplication status of tissues might play an important role, so endopolyploidy was analyzed by flow cytometry in different organs and major differences were revealed. Endoreduplication was minimal in leaves but in other organs—particularly in the hypocotyl and epicotyl—levels up to 64C were found. The basal part of cotyledons has been reported to be the most responsive for adventitious regeneration and, using image cytometry, we identified it as the least endoreduplicated section, with a cycle value of 1.29 compared to 1.78 and 1.80 in the central and distal parts. Basal cotyledonary explants were subjected to various media combinations, revealing Murashige and Skoog (Physiol Plant 15:473–497, 1962, doi:10.1111/j.1399-3054.1962.tb08052.x) medium supplemented with 1 mg L−1N6-benzylaminopurine (BA), 0.25 mg L−1p-aminobenzoic acid (PABA) and 5 mg L−1 fusaric acid (FA) to be optimal (73.3 % regenerated explants with 1.8 shoots per regenerating explants). FA was added to media as a possible selective agent for somaclonal variation inducing increased tolerance to Fusarium but was surprisingly found to stimulate regeneration at a low concentration (5 mg L−1) and to induce genome doubling at medium concentrations (10 and 20 mg L−1).